Part:BBa_K5375001
pA7-GFP
pA7-GFP
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Plasmid lacks a prefix.
Plasmid lacks a suffix. - 12INCOMPATIBLE WITH RFC[12]Plasmid lacks a prefix.
Plasmid lacks a suffix. - 21INCOMPATIBLE WITH RFC[21]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BamHI site found at 4127
Illegal XhoI site found at 3302 - 23INCOMPATIBLE WITH RFC[23]Plasmid lacks a prefix.
Plasmid lacks a suffix. - 25INCOMPATIBLE WITH RFC[25]Plasmid lacks a prefix.
Plasmid lacks a suffix. - 1000INCOMPATIBLE WITH RFC[1000]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BsaI site found at 4091
Contents
Origin
Synthesized by company
Properties
Expression of protein GFP
Usage and Biology
Green Fluorescent Protein (GFP) is a bioluminescent protein initially isolated from the jellyfish *Aequorea victoria*, characterized by its distinctive biological properties and extensive applications. GFP exhibits spontaneous green fluorescence without the necessity for any substrates or cofactors, rendering it an invaluable tool in biological research.
In cell biology and molecular biology, GFP is frequently employed as a reporter gene through fusion with the coding sequence of target proteins, enabling real-time monitoring of their localization, dynamic alterations, and expression patterns within live cells. Furthermore, GFP can be utilized to label specific cellular organelles or structures, thereby assisting researchers in observing cellular processes such as protein transport, signal transduction pathways, and cell division. Owing to its stability and ease of detection, GFP has become an essential component of contemporary life sciences research and has significantly advanced our understanding of cellular biological mechanisms.
Cultivation, Purification and SDS-PAGE
We performed pA7-GFP plasmid linearization by enzyme digest and electrophoresis. We utilized this digest DNA fragment for ligation with the target gene.
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- Figure 1.** The vector of pA7-GFP was enzyme digested.
Measurement and Characterization
To obtain the fusion protein and expression, we constructed plasmids **pA7-HSP70-GFP** and **pA7-PFN3-GFP** and validated them by Sanger sequencing as shown in the figures.
A. **pA7-GFP-PFN3**
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B. **PA7-GFP-HSP70**
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- Figure 2.** Sanger sequencing map of PA7-HSP70 and PA7-PFN3.
Reference
Chalfie M., Tu Y., Euskirchen G., Ward W. W., & Prasher D. C. (1994). Green fluorescent protein as a marker for gene expression. *Science*, 263(5153), 802-805. https://doi.org/10.1126/science.8333281
Cava F., de Pedro M. A., Blas-Galindo E., Waldo G. S., Westblade L. F., & Berenguer J. (2008). Expression and use of superfolder green fluorescent protein at high temperatures in vivo: a tool to study extreme thermophile biology. *Environmental microbiology*, 10(3), 605–613. https://doi.org/10.1111/j.1462-2920.2007.01482.x
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